Poly-gamma-glutamic acid (also known as polyglutamate and PGA) is a biological polymer whose molecular weight can vary anywhere from 20,000 daltons to over 2 million depending on the method of production. PGA is a highly anionic homo-polyamid, whose only components are D- and L-glutamic acid. PGA forms elongated polymer chains through the formation of bonds at the alpha-amino and gamma-carboxylic acid groups. PGA is water soluble, biodegradable, edible and non-toxic toward humans. It is a major component of “natto”, a traditional fermented soybean food in Japan.
PGA was first discovered and reported in 1937 by Ivanovics and co-workers who observed it released upon the cell lysis of Bacillus anthracis. In 1942, Bovarnick reported that Bacillus subtilis secreted PGA into its growth media. Various other Bacillus species were also found to produce PGA externally when fermented. The majority of these findings were published starting in the 1980's and early 1990's.
PGA has a very high negative charge density. The un-ionized form of the molecule adopts the configuration of a helix, whereas the ionized version maintains a random coil configuration.
PGA has been slow to find commercial application. There are several reasons for this. First, it was discovered in a dangerous human pathogen. Second, though glutamic acid is native to humans, poly-alpha-glutamic acid is not. Poly-alpha-glutamic acid is one of two possible isomers and is formed when PGA is manufactured by synthetic means. Poly-alpha-glutamic acid is the most common type of PGA available commercially. Poly-gamma-glutamic acid is the natural form of PGA. It is rare and commercially available from South Korea and Taiwan, but only in low molecular weight and low quality (i.e., not both high molecular weight and medical grade).
Several applications of PGA include environmental/industrial, agricultural, food, and pharmaceutical. One environmental application of PGA is its use as a flocculent. Another newer environmental application of PGA is in removing heavy metal contaminants, such as those used by the plating industry. As mentioned previously, PGA has a very large anionic charge density. Contaminants such as copper, lead, mercury and other positively-charged metal ions associate very strongly with PGA, and can then be concentrated and removed from the waste stream.
Since PGA is comprised of an amino acid, it is an excellent source of nitrogen. This suggests an application in agriculture as a fertilizer. For analogous reasons it is good for drug delivery. A polymer mixture can be packed with nutrients for a particular crop. Once the fertilizer is applied, it has a longer residence time in the soil since the fertilizer nutrients are protected from the natural environment by the PGA.
In the food industry, work has been done that shows PGA functions as a cryoprotectant. PGA has been shown to have antifreeze activity significantly higher than glucose, a common cryoprotectant. It has also been used as a stabilizer in ice cream and as a thickener in juice.
In the medical field, PGA is being studied as a biological adhesive and a drug delivery system. Gelatine-PGA solutions, and cross-linked PGA solutions have shown application as adhesives without the toxic or inflammatory issues. PGA has also been used in drug delivery. Taxol®, a well know cancer drug by Bristol-Myers Squibb, was covalently linked to PGA. The resulting molecule, (PG-TXL) in pre-clinical testing, showed a five-fold increase in tumor uptake of Taxol®.
Large MW PGA has advantages over low MW PGA including higher charge densities and higher viscosities at lower concentrations. This means that high molecular weight PGA would have advantages, including (1) greater reactivity with alkaline materials to make soap and other consumer products, of which the high viscosity is a required property, (2) more nitrogen delivered making it very useful in agriculture, (3) higher drug loading at the active negative sites, and (4) higher viscosities resulting in better drug diffusion properties.
The ability to deliver high MW PGA of the correct purity for the application is key.
Lastly, there is the issue of molecular weight and how it is measured. Several groups claim to have or produce high molecular weight gamma isomer PGA (claims range from 1 to 4 million). In general, these groups are using analytical methods not suited to PGA analysis. Most groups utilize size exclusion chromatography at neutral pH and physiological ionic strength. Under these conditions, PGA interacts with commercial columns, shifting peak retention times and giving erroneous results. In addition, these retention times must be compared to standards, which are typically non-ionic polymers. These standards do not have the same radius of gyration and thus do not behave like PGA, therefore results are typically incorrect.
In order to properly determine the molecular weight of PGA, one may employ an analytical method that involves low pH and low ionic strength, and couples size exclusion chromatography with multi-angle laser scattering, as described in a Master's Thesis By Louis R. Stock II entitled “Rheological Characterization of (Poly-γ) Glutamic Acid Fermentations” (1996) (incorporated herein by reference). Under these conditions, with the anionic sites fully protonated, PGA molecular weights may be correctly determined. Analysis under these conditions has established that the molecular weights of samples reported to be 1-4 million are in fact 25,000 to 400,000. There is thus a need for an economical, practical method of producing PGA at both low and high molecular weights with purities appropriate for both human and non-human uses.